Tuberculosis (TB) has been increasing in incidence over the last seven years. Of particular concern is the marked increase in multidrug resistant (MDR)-Tb. In 1991, Bellevue Hospital treated 506 new cases of Tb, 80% of these patients were African American or Hispanic and 16% had MDR-Tb. This epidemic is being investigated using restriction fragment polymorphisms (RFLP). Of 289 clinical isolates examined, 39 different patterns named A through MM have been identified. Different RFLP types have different incidence of MDR, ranging from none of eighteen in strain C to ten of ten in strain W. The high molecular weight DNA separation technique of pulse field gel electrophoresis will be used to develop a new RFLP system which identifies drug resistant strains, defining close genetic linkage of drug resistance with RFLP patterns. Another series of studies will search for plasmid mediated drug resistance in M. tuberculosis (M.tb.) using sucrose gradient centrifugation to separate plasmids from chromosomal DNA and identify exchange of drug resistance genes from one strain of M.tb. to another strain by cocultivation and subsequent drug selection. The goal of this investigation is to better ,understand the mechanisms which generate drug resistance in M.tb. and to clone drug resistance genes. Once an element has been identified in the above investigations, we will use this putative drug resistance gene to confer drug resistance to a sensitive M.tb. strain, thereby proving that the element is a drug resistance gene. The molecular tools generated will allow more rapid diagnosis of MDR-Tb and early institution of effective antibiotic therapy. The understanding of generation of MDR-M.tb. will be valuable in the study of the epidemiology of MDR-Tb and contribute to generating effective Public Health policy for controlling this devastating infection.